The purpose of this project is to characterize the nuclear binding of the estrogen receptor and to evaluate the control of recycling and/or synthesis (replenishment) of the cytoplasmic receptor. Nuclear binding of the estrogen receptor complex to specific acceptor sites and non-specific sites will be evaluated by differential salt extraction and by kinetic analysis. The various forms of the receptor will be examined during nuclear residence and the mechanisms by which the receptor estrogen complex dissociated from nuclear sites will be studied. The replenishment of the cytoplasmic receptor subsequent to nuclear binding will be examined by using the differential effects of estriol, progesterone and non-steroidal estrogen antagonists on the replenishment process. The mechanisms by which estrogen antagonists block or alter the nuclear binding of the receptor estrogen complex will be examined. The influence of estrogen antagonists on RNA polymerase activity and the number of polymerase initiation sites on chromatin will be examined in relation to estrogen receptor binding and processing in the nucleus. The 3H-steroid exchange assay will be further characterized and extended for use with other steroid hormones. This will be done by a systematic analysis of the various parameters that influence the exchange process, i.e., temperature, time, pH, ionic strength, etc. The receptor estrogen-nuclear binding will also be characterized as the exchange process takes place in an attempt to relate these reactions to those that occur in vivo.